Nanostructure-Ended Closed Linear DNA for Scalable In Vivo Production

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Solution Overview

Problem

Current methods for producing nanostructure-ended double-stranded covalently-closed linear DNA molecules face challenges in scalability and cost-effectiveness, limiting their application in medical and industrial fields.

Innovation Solution

A novel in vivo manufacturing system utilizing a recombinant cell with a Parental Plasmid DNA Platform, comprising Retron, Linear, and Bacterial Backbone modules, along with specific promoters and enzymes, to synthesize nanostructure-ended double-stranded covalently-closed linear DNA molecules, enabling customizable DNA ends and a genetic expression unit.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If traditional methods are used to produce nanostructure-ended double-stranded covalently-closed linear DNA molecules, then production is possible, but scalability and cost-effectiveness are limited

Engineering Contradiction:

Improveproduction scalabilityVSAvoidcost-effectiveness

Core Design Contradiction:

ProductivityVSEase of manufacture

Solution Approach 1:

The system divides the DNA manufacturing process into modular components housed within a recombinant cell: a Repressor Protein Module, a Recombinase-Reverse Transcriptase Module, and a Homing Endonuclease Module, each controlled by specific promoters and ribosome binding sites. This segmentation allows independent optimization and scalable production of each functional module.

Inventive Principle:

Principle #1Segmentation

Solution Approach 2:

The parental plasmid DNA platform serves multiple functions simultaneously: it houses retron units for generating DNA nanostructures, contains linear modules for assembling the final product, and includes bacterial backbone elements for replication and selection. This multi-functionality consolidates what would otherwise require multiple separate manufacturing steps into a single integrated system.

Inventive Principle:

Principle #6Universality (Multi-functionality)

2Adaptability or versatility

If complex three-dimensional structures are created to achieve protein-like functions, then functional versatility is improved, but stability deteriorates due to environmental sensitivity

Engineering Contradiction:

Improvefunctional versatilityVSAvoidstructural stability

Core Design Contradiction:

Adaptability or versatilityVSStability of the object's composition

Solution Approach 1:

The system uses DNA nanostructures as stable copies or substitutes for protein structures. The retron units generate DNA sequences that fold into protein-mimicking three-dimensional configurations, providing the functional versatility of proteins while maintaining the chemical stability and environmental resistance inherent to nucleic acids.

Inventive Principle:

Principle #26Copying

Solution Approach 2:

The system creates hybrid functional elements by combining DNA nanostructures with peptide sequences through the double-stranded linear DNA linker. This composite approach allows the DNA portion to provide structural stability while the peptide portions contribute protein-like functional capabilities, achieving both stability and versatility simultaneously.

Inventive Principle:

Principle #40Composite materials

Applied Scientific Principles

This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.

Function Achieved in This Case

The system provides scalable and cost-effective production of DNA molecules with protein-like functions, suitable for various medical and industrial applications, including gene therapy and immunotherapy.

Implementation Method 1

a Recombinase-Reverse Transcriptase Module comprising: a First Bacterial Inducible Promoter, a Third Ribosome Binding Site, a Recombinase Coding Sequence, a Fourth Ribosome Binding Site, a Reverse Transcriptase Coding Sequence, and a Second Bacterial Terminator

Methodology Applied

Scientific EffectReverse transcription:

Implementation Method 2

a Recombinase-Reverse Transcriptase Module comprising: a First Bacterial Inducible Promoter, a Third Ribosome Binding Site, a Recombinase Coding Sequence

Methodology Applied

Scientific EffectSite-specific recombination:

Implementation Method 3

a Homing Endonuclease Module comprising: a Second Bacterial Inducible Promoter, a Fifth Ribosome Binding Site, a Homing Endonuclease Coding Sequence, and a Third Bacterial Terminator

Methodology Applied

Scientific EffectHoming endonuclease cleavage:

Implementation Method 4

a First Bacterial Constitutive Promoter, a First Ribosome Binding Site, a First Repressor Protein Coding sequence

Methodology Applied

Scientific EffectTranscription:

Implementation Method 5

a First Ribosome Binding Site, a First Repressor Protein Coding sequence

Methodology Applied

Scientific EffectTranslation:

Data Source

PatentUS12509711B1System and process for in vivo manufacturing nanostructure-ended double-stranded covalently-closed linear DNA, the resulting molecules and their uses

Publication Date: 2025.12.30 SYSTE BIO INC

AI summary

The present invention relates to a novel biological in vivo manufacturing system and a process for generating nanostructure-ended double-stranded covalently-closed linear DNA molecules. These DNA molecules possess the ability to merge the information-storage and function-encoding attributes of nucleic acids with the structural properties and functional capabilities typically found in proteins, such as specific binding and catalysis, in a single nucleic acid-only molecular entity, making them useful for a wide variety of medical applications and industrial implementations.